The proposal will investigate two problems of aging as they relate to connective tissue metabolism. During the past funding period findings from this program have held considerable influence in the field of wound healing. Using rat models that reflect normal healing, the diabetic state, and aging, we have either published or presented data showing how the process and defects of tissue repair can be modulated by the availability of cellular growth factors. Further work is proposed to determine which factors are critical to the healing process by four approaches: (a) addition of growth factors alone, in combination, or in sequence to achieve quantitative reversal of healing defects in age-impaired wound healing; (b) introduction of growth factor-encoding sequences into wound sites by transient in vivo transfection of eukaryotic expression vectors or mRNA using cationic liposomes; (c) use of neutralizing antisera and antisense vectors or oligonucleotides at the wound site to identify critical cytokine pathways; (d) in situ hybridization to quantify responses of matrix genes. Principal matrix molecules to be evaluated at the RNA level include collagen types I and III, elastin, the metalloproteinases stromelysin and collagenase, their inhibitor, TIMP, and several cytokine genes. Key agents will be those available in quantities sufficient for animal studies: transforming growth factor-alpha, transforming growth factor-beta-1 (TGF- beta-1), basic fibroblast growth factor (bFGF) and platelet-derived growth factor. Our aim is to provide closer correlation between the biological effects which can be observed in the animal models and those effects observed in isolated cell culture. Premature aging syndromes are thought to exaggerate the normal process of aging. Elastin loss and degradation are familiar manifestations of cutaneous aging. In addition, elastin production appears to serve as a useful marker of certain forms of premature aging in fibroblast strains, possibly reflecting an altered response to cytokines. The second area of investigation focuses on the cellular behavior of a set of segmental aging syndromes, Hutchinson-Gilford progeria, Werner's syndrome, and Cockayne syndrome, each of which show unusual responses to the cytokines, TGF-beta and bFGF, in terms of their expression of the matrix protein, elastin. Experiments are proposed to determine if such cells overexpress trans-acting factors that impinge on the elastin promoter and if they secrete autocrine factors that affect elastin production by normal cells.